Chelated 8-hydroxyquinoline and use thereof in a method of treating epithelial lesions

ABSTRACT

A chelation complex including 8-hydroxyquinoline and zinc mixed with a carrier demonstrates therapeutic efficacy in treating lesions including cancerous lesions, precancerous lesions, cysts and warts.

RELATED APPLICATIONS

This application is a divisional of application Ser. No. 10/247,161filed Sep. 18, 2002 now U.S. Pat. No. 7,060,696 which is a divisional ofapplication Ser. No. 09/601,304 filed Jan. 2, 2001 now U.S. Pat. No.6,476,014 claiming benefit of priority to PCT/US99/02817 published onAug. 12, 1999 and is a continuation-in-part of application Ser. No.09/021,421 filed Feb. 10, 1998 now U.S. Pat. No. 7,846,919.

BACKGROUND OF THE INVENTION

1. Field of the Invention

The present invention pertains to the field of oxinates and,particularly, to the therapeutic use of metal chelated8-hydroxyquinolinates in the treatment of cancers, precancerous lesions,and other abnormal tissues.

2. Statement of the Problem

A preferred treatment modality for many cancers is surgical excision ofthe cancerous lesion. Surgical excision is not always desirable whensurgery could sever nerves or produce scars that interfere with normalmovements in tissues proximate the site of surgery.

Chemicals have been developed to treat cancers. Rate-sensitive cytotoxicdrugs have cytotoxic effects on all tissue types, but are particularlyeffective against certain cancers that function at a metabolic rategreater than the metabolic rate of normal cells. The increased metabolicrate of these cancers makes them more susceptible to the cytotoxicity ofthe drugs. In this manner it is possible to provide a dosage that isfatal to cancer cells, while that same dosage is not fatal to normalcells. By way of example, U.S. Pat. No. 5,684,169 teaches the formationof cyclodextrin complexes of taxol to improve the solubility of taxol inwater. Taxol is a cytotoxic drug that is believed to attack and killcancer cells with a rate-sensitive effect.

Other chemicals are specifically designed for the treatment of skincancers and cancers that reside close to the skin. U.S. Pat. No.5,605,700 describes a transdermal preparation containing toremifene foruse in treating cancers of the skin or cancers that reside a shortdistance from the skin, such as metastic lesions of breast cancer. Thetransdermal preparations are said to be of particular interest in thetreatment of melanoma, lymphoma, Kaposi's sarcoma, and fungoidesmycosis. Escharotics or caustic chemicals, such as Podophylin ortricholoroacetic acid which are used in the treatment of skin cancer,venereal warts and human papilloma virus, are designed to produce achemical burn that destroys substantially all of the tissues in contactwith the chemical. None of these chemicals is selective in destroyingonly the lesion while leaving behind healthy tissues that exist close tothe site of the lesion, i.e., the chemicals lack selectivity andspecificity.

The chemical 8-hydroxyquinoline is not used for the treatment ofepithelial lesions, such as epitheliomas and venereal warts.8-hydroxyquinoline is known by various other names includingoxyquinoline, oxine, 8-hydroxy-chinolin, hydroxybenzopyridine, and8-oxyquiniline. The federal government has approved 8-hydroxyquinolinefor cosmetic use in low concentrations of less than five percent. Atthese concentrations, 8-hydroxyquinoline functions as a cosmeticbiocide, as reported in JACT 11(4), 1992. U.S. Pat. No. 4,302,467reports the use of 8-hydroxyquinoline or its chelates in combinationwith dehydroacetic acid or sorbic acid. The combination is reported towork synergistically against bacteria and fungus. According to the MerckIndex 11th Edition, Merck & Co. publ., p. 4779. (1989),8-hydroxyquinoline is used as a fungistat, a chelating agent in thedetermination of trace metals, and a disinfectant.

Chapter ten of the book Albert, Selective Toxicity 3rd Ed., New York,John Wiley & Sons, Inc. (1965) states on page 370-378 that theantibacterial action of oxine is due to chelation. Iron-chelated formsof oxine are toxic to gram-positive bacteria when mixed with Fe³⁺ at a1:1 molar ratio and a 1:2 molar ratio of oxine to iron, but toxicitydiminishes at a 1:3 ratio. Despite its strong antibacteriologicaleffects, oxine and its derivatives are not normally injected into thebloodstream because they are inactivated by red blood cells, whichsecrete a substance that binds oxine.

There remains a need for a topically applied therapeutic compositionwith selective toxicity in the treatment of epithelial lesions.

SOLUTION

The present invention overcomes the problems that are outlined above byproviding a topically applied therapeutic composition having selectivetoxicity in the treatment of epithelial lesions. The therapeuticcomposition demonstrates selective toxicity on human lung cancer, breastcancer, and melanoma. Furthermore, the composition has been used totreat human patients where it demonstrates a one-hundred percenttherapeutic index with selective toxicity against venereal warts, maleveruoca warts, lesions produced by human papilloma virus, basal cellcarcinoma, solar keratosis, and Kaposi's sarcoma. In veterinaryapplications where dogs, cats, and horses are the patients, thecomposition shows a one-hundred percent therapeutic index with selectivetoxicity against eye cancer, sarcoids, sarcoma, malignant melanoma,rectal adenoma, histiocytoma, and sebaceous adenoma. In this context,the therapeutic index is defined as the number of patients cured dividedby the number of patients treated. While chemical excision of theselesions is accomplished, the composition is essentially withoutescharotic effects on healthy tissue.

A therapeutic composition according to the present invention contains8-hydroxyquinoline or a functional homologue thereof, a metal chelatingagent, and a carrier. The 8-hydroxyquinoline and the metal chelatingagent are present in effective amounts for treating mammalian epitheliallesions selected from the group consisting of cancerous lesions,precancerous lesions, and warts. These mammalian epithelial lesionsspecifically include venereal warts, male veruoca warts, lesionsproduced by human papilloma virus, basal cell carcinoma, solarkeratosis, Kaposi's sarcoma, eye cancer, sarcoids, sarcoma, malignantmelanoma, rectal adenoma, histiocytoma, and sebaceous adenoma.

In veterinary applications, solutions according to the present inventionhave been used with success to treat dermal lesions including acanthosisnigricans, acne, acral lick dermatitis, allergic reactions, calcinosiscircumscripta, calcinosis cutis, cutaneous asthenia, deep mycoticinfection, demodicosis, acute and chronic dermatitis, dermatomycosis,eosinophillic granuloma complex, epidermal cysts, hypothyroidism,ichthyosis, insect bites, lentigo, nodular penniculitis, pemphigus,canine scabies, and thallium toxicosis. The solutions have also beenused with success to treat veterinary tumors including basal celltumors, fibroma, fibrosarcoma, granuloma, hemangioma,hemangiopericytioma, histiocytoma, intracuaneous cornyfying epithelioma,lipoma, lymphosarcoma, mast cell tumor, melanoma, papilloma, perianaadenoma, reticulum cell sarcvoma, sebaceouos gland tumor, squamous cellcarcinoma, sweat gland (apocrine) tumor, and transmissible venerealtumor. Injectable forms of the solutions have been used to counteractthe effects of bites from the brown recluse spider, which can havedisfiguring and painful consequences if left untreated.

Therapeutically effective amounts include therapeutically effectiveconcentrations, as well as therapeutically effective ratios of the8-hydroxyquinoline and the metal chelating agent. The preferred rangefor therapeutically effective ratio is one wherein the molar ratio of8-hydroxyquinoline to the metal chelating agent ranges from 1:1 up to 1:N, where N is the oxidation state of the metal chelating agent. At theratio of 1: N, the chelated oxine is easily transported across the cellmembrane because it has a neutral charge. Charged species are alsoobserved to be threrapeutically effective. Where the metal has a valenceof +2, therapeutically effective ratios range from 1:1 to 1:3, and themost preferred ratio is about 1:2. The 8-hydroxyquinoline is preferablypresent in an amount ranging from five percent to twenty percent of thecomposition by weight, and the most preferred amount of8-hydroxyquinoline is about ten percent by weight. More or less8-hydroxyquinoline may be used. For example, injectable forms or orallyconsumed forms may have reduced concentrations for long term uses, suchas one half percent to five percent concentrations of chelated8-hydroxyquinoline to be used as continuing maintenance treatments for acancer patient who is in remission but remains at risk for a recurrenceof cancer. The amount of 8-hydroxyquinoline may be increased beyondabout twenty percent by weight, but these higher concentrations, incombination with zinc chloride chelating agent, are likely to induce asensitized reaction in the patient. Use of these higher concentrationsmay be justified when faced with a tumor or lesion that is particularlydifficult to remove.

As stated above, the therapeutic compositions include a metal chelatingagent. This agent is preferably a chelatable metal bonded to a halogen,which yields an ionic species of the metal in solution. This reaction isfree for covalent and coordinate bonding with oxine in solution. Themetal is preferably a heavy metal or a transition metal. Heavy metalsare more preferred, and the preferred heavy metals are copper, iron,manganese, molybdenum, and cobalt. Zinc is the most preferred heavymetal. The metal is preferably provided as a salt including a metal anda halogen, thought he halogen is not required for purposes of theinvention. Chlorine is the most preferred halogen. Thus, the mostpreferred metal chelating agent is provided as zinc chloride.

The carrier is preferably formed of a gel base to enhance the timeduring which the composition is retained on the epithelial lesion whereit is applied. Lecithin is preferably added as a penetrating agent, andthe penetrating effect of lecithin may be enhanced by the use ofdimethyl sulfoxide (DMSO). The carrier also contains an emollient, suchas isopropyl palmitate or isopropyl myristate. The gel base is made of ahydrophilic polymer, such as polyacrylamide or polyoxyalkylenederivatives of propylene glycol. A topical steroid, such as 1-2%hydrocortizone may be added to decrease the incidence or severity ofcontact dermatitis proximate the site of topical application.Humectants, such as propylene glycol may be added to reduce dryness ofthe lesion during the course of treatment.

A further preferred aspect of the invention is the use of antioxidantsto stabilize chelated 8-hydroxyquinoline in vivo. Nordihydroguiareticacid, derivatives of nordihydroguiaretic acid, and functional homologuesof nordihydroguiaretic acid stabilize zinc-chelated 8-hydroxyquinolineagainst inactivation by thermolabile substances in the blood forsolutions that are injected into the body. Similarly, 8-hydroxyquinolineand zinc chloride coprecipitated with sodium ascorbate forms a stableorally administrable product having therapeutic efficacy.

The composition is administered to the epithelial lesion by topicalapplication, injection, or oral administration. Where the lesion isscarred or thickening is observed, topical efficacy is enhanced byscraping and perforating the lesion with a needle or scalpel prior totopical application of the ointment, in order to enhance the penetrationof the ointment into the lesion. Only one topical application istypically required, and the tissue forming the lesion is expected toturn black and necrotic after about two to four weeks. The dead tissueis peeled away, and the wound site typically heals to completion afteranother three to four weeks have passed. Even where large lesions havebeen removed in this manner, there is typically no scarring, and eventhe hair follicles are restored to full function. Additional amounts ofointment may be applied if tumor necrosis is not proceeding quicklyenough. Additional solution is preferably applied to the wound marginsafter the removal of necrotic tissue to assure that all of the cancercells have been eliminated.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 depicts the chelation mechanism between a metal and oxine.

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENT

FIG. 1 depicts the chelation mechanism for 8-hydroxyquinoline. A metalchelating agent M 100 has an oxidation state of N+. The metal chelatingagent M 100 reacts with 8-hydroxyquinoline 104 to produce a 1:1 metaloxinate 104 having a valence of N-1. A second 8-hydroxyquinolinemolecule 106 reacts with the 1:1 metal oxinate 104 to produce a 1:2metal oxinate 108. This process is repeated N times until the resultantoxinate has a neutral charge. even then there is coordinate attractionbetween the metal M 100 and the heterocyclic nitrogen atoms in otheroxinates. The lowest ratio oxinates, e.g., the 1:1 oxinate 104, are themost stable products due to the steric hindrance that the large oxinateligands provide as the reaction proceeds. Thus, if the reaction islimited by the 8-hydroxyquinoline reagent, the lower ratio species willpredominate. Oxine has a high avidity for metals, and the reaction goesto substantial completion.

Ionic reaction products, i.e, those having a ratio less than 1: N, aresubjected to partition effects where cell membranes regulate the intakeof cations. On the other hand, uncharged complexes are liposoluble, andreadily penetrate cell membranes in a manner in which the cell cannotregulate. While 8-hydroxyquinoline is normally not toxic, chelated formsof 8-hydroxyquinoline do have cytotoxic effects in mammalian cells. Thiscytotoxic effect is empirically observed for both the charged anduncharged reaction products.

A general theory of pharmacokinetics in the invention is that abnormalmammalian epithelial cells have membranes which are more permeable tochelated forms of 8-hydroxyquinoline than are normal cell membranes.Thus, the chelated 8-hydroxyquinoline is transported across abnormalcell membranes in a greater concentration than is permitted by normalcell membranes. It follows that topically applied, orally administeredor injected forms of chelated 8-hydroxyquinoline have a greatercytotoxic effect upon the abnormal cells because abnormal cells absorbit in a greater concentration. This theory accounts for the observedselective toxicity to epidermal lesions and cancerous tumors. It doesnot necessarily matter whether the cytotoxicity inside the cell derivesfrom the metal chelating agent or the 8-hydroxyquinoline, but one theoryof operation explaining the observed cytotoxicity is that theoxyquinoline is the transport agent and the metal chelating agent,preferably, a heavy metal chelating agent, is primarily responsible forthe observed cytotoxicity inside the cell. Thus, it is possible toincrease the selective or specific cytotoxic effects of the chelatedcompound by increasing the toxicity of the heavy metal, e.g., byselecting osmium or arsenic. The metal may also be a radioactive isotopehaving a short half life of a few days or hours, such as ²⁸Mg, ¹⁹⁸Au,and ⁵²Fe.

In addition to topical applications of the compositions,nordihydroguiaretic acid is used to stabilize the chelated8-hydroxyquinoline in solutions that are injected into the patient.Nordihydroguiaretic acid is an antioxidant that protects the chelatedoxine compositions against inactivation that, otherwise, might occur dueto secretions from the blood that tend to inactivate oxine. Similarly,another antioxidant, sodium ascorbate, is coprecipitated with oxine andzinc chloride from a solution of ethanol and water to form an orallyadministrable product, which also has in vivo utility.

Patients sometimes report a burning sensation at the lesion aftertopical application of the chelated oxinates. Therefore, it is preferredto place an emollient into the composition, in order to sooth theburning sensation. Penetrating agents, such as lecithin or DMSO, enhancethe efficacy of the compositions.

The following working examples set forth preferred methods and materialsfor use in practicing the invention.

EXAMPLE 1 Preparation of a Zinc-Chelated Liquid Solution of8-Hydroxyquinoline for Use in Treating Epithelial Lesions

Bulk stock solutions were made for later use as intermediates in mixingthe final solution. The bulk stock solutions included a gel basesolution that was used to stabilize the final product and impart asuitable thickness for the intended environment of use. Alecithin/isopropyl palmitate solution was also prepared.

The gel base solution was prepared by mixing 20 grams (gm) of PluronicF127™¹ with 0.2 gm potassium sorbate. Purified water was added to bringthe total volume to 100 ml. Pluronic F127™¹ is a high molecular weightpolyoxyalkylene ether derivative of propylene glycol having watersoluble, surface active, and wetting properties. The ingredients weremixed in a Braun mixer, sealed in a bowl, and stored under refrigerationat about 5° C. once all of the granules became wet. The solution wasrefrigerated to avoid solidification of the gel, which occurs at roomtemperature. ¹ Pluronic F127 is a trademark of the BASF Corporationlocated in Parisippany, New Jersey.

The lecithin/isopropyl palmitate solution was prepared by dispersing 100gm of granular soya lecithin and 0.66 gm of sorbic acid in isopropylpalmitate (100 gm or 117 ml). The dispersed mixture was allowed to standovernight at room temperature of about 20° C. A liquid formed having asyrup-like consistency.

A 100 gm aliquot of the gel base solution was mixed with 22 ml of thelecithin/isopropyl palmitate solution and 10 ml of distilled water. Themixture was stirred to visual homogeneity. Ten gm of 8-hydroxyquinolineand 20 gm of ZnCl₂ were added to the mixture, which was again stirred tohomogeneity.

EXAMPLE 2 Topical Application of a Zinc-Chelated 8-HydroxyquinolineSolution to Cure Athymic Nude Mice of Cancerous Lesions Corresponding toExplanted Samples of Human Lung Cancer

A live human lung cancer cell line Calu-1 or SW-900, as reported byFogh, J., et al. 59 J. Natl. Cancer Inst. 221-225 (1977), is an epidurallung cancer having an undifferentiated in vitro cytopathology. Celllines of this type are known to grow tumors of correspondingcytopathology when explanted into athymic nude mice, as reported by Foghand Giovanella eds., The Nude Mouse in Experimental and ClinicalResearch, Academic Press, London, pp 235-266 (1978) (see especially p.239). Athymic nude mice may be purchased from specially maintainedbreeding colonies throughout the United States, such as the breedingcolony at the University of Nevada in Reno, Nev. Athymic nude mice areespecially desirable to test the efficacy of anticancer compositionsbecause athymic mice lack immune systems. Thus, the mice do not haveimmune systems to attack explanted cancers in the mice, and the efficacyof anticancer compositions upon such cancers is primarily due to thecompositions alone.

The test procedures that are outlined below are substantially the sameas procedures that have actually been performed on athymic nude mice.The procedures below have been modified slightly to present the mostpreferred mode of performing the procedures.

One hundred and fifty athymic nude mice are obtained from a reputablesupplier, e.g., the breeding colony at the Worchester Institute, inWorchester, Mass. The mice are split into a treatment test population offifty mice, a no treatment test population of fifty mice, and a controlpopulation of fifty mice. The populations are maintained as separategroups using conventional procedures for maintaining breeding coloniesof athymic nude mice under sterile conditions.

A human epithelial lung cancer cell line, e.g., the Calu-1, SW-900, orMX-1 lung adenocarcinoma line, is cultured in a suitable growth medium,e.g., 50 ml of Eagle's MEM medium (purchased from Life Technologies,Inc., Gaithersburg, Md.), which is supplemented with 10% fetal calfserum and 50 μg/ml gentamicin from Sigma Chemical Co. of St. Louis, Mo.The cells are maintained in a 5% humidified CO₂ atmosphere at 37° C. in75 cm² plastic tissue culture flasks, with media passage at 5-7 dayintervals.

A sample of approximately five million cancer cells is taken by 20 gaugetrocar and extruded into the subcutaneous tissue at the posterior baseof the neck on an athymic nude mouse. The procedure is repeated for eachmember of the treatment and no treatment test populations. Conventionalsterile procedures are used while working with the mice to avoidinfecting the mice with pathogens other than the cancer cell line. Livetissue from surgical patients having human lung cancer may besubstituted for the cultured cell lines.

Approximately one-hundred percent of the mice can be expected to survivethe grafting procedure. If proper explantation procedures are used, aswill be understood by those who are skilled in those procedures,approximately one-hundred percent of the explanted cancer cells willproduce epithelial lesions proximate the site of trocar insertion intothe mouse. The explanted cancer is permitted to grow for approximatelytwo to three weeks until a cancerous epithelial lesion of approximately1.5 to 2 cm in diameter is externally visible.

The untreated test population will typically die within sixty days. Oncedeaths begin occurring, which is usually within about thirty days, it ispreferred to euthanize the entire untreated test population to preventunnecessary suffering. The deaths typically occur from strangulation dueto growth of the cancerous tumor at the base of the neck.

The topical solution that was produced in Example 1 is removed fromrefrigerated storage and stirred to substantial homogeneity. A cottonswab is dipped into the solution. An athymic nude mouse having aposterior neck lesion approximately 1.5 to 2 cm in diameter is selectedfrom the treatment test population. A needle is used to perforate theepidermal portion of the lesion at locations spaced about one millimeterapart from one another. A swab is used to apply a coating of solutionover the entire lesion and extending at least one-half cm beyond thevisible margins of the lesion. This procedure is repeated for eachmember of the treatment test population. A single topical application ofthe solution is the sole and exclusive time that the solution is appliedto the lesions.

The treatment test population is monitored for nine months after topicalapplication of the solution. Notes are taken as to the progress ofcancer in the mice. The lesions appear inflamed on the first day aftertopical application. Three to five days after topical application, thelesions appear black with white streaks on the surface. Eight to tendays after the topical application, the lesions appear black andnecrotic. Fourteen to sixteen days after the topical application, thelesions have sloughed off. Pathologic examination of tissue samplestaken from ten individual mice at the interior margins of the sloughedlesion confirms that no cancer is visibly apparent at the margins. Inthee to four weeks after topical application, the tissue at the formerlesion sites has healed to become substantially indistinguishable fromnormal tissue with no evidence of scarring. The treatment testpopulation exhibits cachexia or a slight emaciation corresponding to anaverage weight of about 4 to 5 gm less than the control population.

The treatment test population mice and the control population mice aremonitored for nine months after topical application of the solution.One-hundred percent of the treatment test population typically survivesthe procedure with no evidence of cancer after nine months, i.e., thetherapeutic index of the solution approximates one hundred percent. Thissuccess is contrasted against a one hundred percent mortality in theuntreated test population, if the untreated test population has not beeneuthanized to prevent unwarranted suffering.

EXAMPLE 3 Topical Application of a Zinc-Chelated 8-HydroxyquinolineSolution to Cure Athymic Nude Mice of Cancerous Lesions Corresponding toExplanted Samples of Other Human Cancers

The procedure of Example 2 is repeated for other cancers including humanbreast cancer, human colon cancer, and human melanoma. It is noted thatlung, breast, and colon cancers are classified as malignant epithelialtumors, while melanoma is a malignant nonepithelial tumor. Thus, thestudy addresses both epithelial and nonepithelial tumors. Specific celllines that may be used include SW-613 (breast duct adenoca), SW-620(colon adenoca), and RPMI-7262 (melanoma). See, for a discussion ofthese cell lines, Fogh, J., et al. 59 J. Natl. Cancer Inst. 221-225(1977), and Fogh and Giovanella eds., The Nude Mouse in Experimental andClinical Research, Academic Press, London, pp 235-266 (1978) (seeespecially pp. 238-239).

The therapeutic index is typically one hundred percent effectivenessagainst all of these explanted cancers in nude mice, with one hundredpercent survivorship through treatment, selectivity for the destructionof diseased tissue alone, and no apparent scarring at the former lesionsite. The cancers showed no sign of recurrence in the treated testpopulations after nine months.

EXAMPLE 4 Preparation of Zinc-Chelated 8-Hydroxyquinoline Solutions forInjection into Athymic Nude Mice

These ingredients are weighed out and combined under sterile conditions:nordihydroguiaretic acid (30 gm), 3% saline in water (30 gm),8-hydroxyquinoline (10 gm), and ZnCl₂ (20 gm). The ingredients arestirred to substantial homogeneity, and stored under refrigeration atabout 5° C. to provide an injectable solution.

Cruder forms of injectable solutions are provided by combining thesolutions of Table 1 with forty to ninety percent by weight alcohol ornormal saline solution. Isopropyl alcohol is particularly preferred foruse in these solutions. Alternatively, zinc chelated oxyquinoline may bemixed directly with alcohol to make a crude injectable solution.

EXAMPLE 5 Injection of Zinc-Chelated 8-Hydroxyquinoline Solution to CureAthymic Nude Mice of Human Cancer Lesions

The studies of Examples 2 and 3 are repeated in an identical manner,except the injectable nordihydroguiaretic acid solution or alcohol basedsolution of Example 4 is substituted for the topical solution ofExample 1. In each case, a one-half ml quantity of the Example 4solution is injected directly into subcutaneous tissue proximate thelesion.

The therapeutic index of the injected solution is typically one-hundredpercent effectiveness against all of the explanted lung, breast, colon,and melanoma cancers in athymic nude mice, with one hundred percentsurvivorship through treatment, and no apparent scarring at the formerlesion site. The cancers showed no sign of recurrence in the treatedtest populations after nine months. The therapeutic effects are superiorwith necrotic effects being observed at the lesion site approximatelythree days sooner than in the case of the topical ointment.

EXAMPLE 6 Various Formulations of Chelated 8-Hydroxyquinoline Solutionsfor Use in the Treatment of Cancerous and Precancerous EpithelialLesions

Table 1 below shows various topical product formulations that may beused to dispense therapeutically effective amounts of 8-hydroxyquinolineand zinc chloride. In Table 1 below, “NDGA” representsnordihydroguiaretic acid. “P20%” represents a solution that contains 20%Pluronic 127 NF¹ by weight, in addition to 0.2% potassium sorbate, and79.8% distilled water. “P30%” represents a solution that contains 30%Pluronic 127 NF™¹ by weight, in addition to 0.2% potassium sorbate, and69.8% distilled water. “P40%” represents a solution that contains 40%Pluronic 127 NF¹ by weight, in addition to 0.2% potassium sorbate, and59.8% distilled water. “Lec/Is” refers to a solution that contains 100gm granular soya lecithin, 100 gm isopropyl palmitate, and 0.66 gmsorbic acid.

The Pluronic 127 NF¹ solutions have pharmacoactivity that facilitatesthe transportation of large molecules through epidermal tissues. In thismanner, the gels function as an adjuvant to improve the efficacy ofchelated 8-hydroxyquinoline compositions. A disadvantage of Pluronic 127NF¹ gels is that they must be made and stored at temperatures less than40° F. because the gels solidify at higher temperatures. Other gel basesmay be used including Aquabase or other polymers dispersed in a suitablesolvent for 8-hydroxyquinoline. Lecithin and other penetrants includingdimethyl sulfoxide may also be used within prescribed limits to enhancethe activity of the chelated oxyquinoline compositions. In Table 1, theterm “q.s.” is used to show that a sufficient quantity of gel was addedto bring the total weight of the combined ingredients to the weightshown in the column, i.e., 100 gm. ¹ Pluronic F127 is a trademark of theBASF Corporation located in Parisippany, N.J.

Other commercially available gel bases are also useful when usedaccording to the manufacturer's instructions. Examples of these gelbases include Aquabase™², which is a hydrophilic combination ofpetrolatum, mineral oil, mineral wax, woolwax, alcohol and cholesterol,and Plastibase 50-W™³. ² Aquabase is a trademark of the PaddockLaboratories located in Minneapolis, Minn.³ Plastibase W-50 is aregistered trademark of E.R. Squibb & Sons located in Princeton, N.J.

TABLE 1 TOPICAL OINTMENT PRODUCT FORMULATIONS 8- Aqua- Plasti- Hydroxy-Zinc Distilled P20% P30% P40% base base Quer- Methyl Sam- quinolineChloride Water NDGA (q.s. (q.s. (q.s (q.s (q.s cetin Larrea Lec/ls DMSOSb₂S₃ Cellulose ple (gm) (gm) (ml) (gm) gm) gm) gm) gm) gm) (gm) (gm)(ml) (ml) (gm) (gm) A — 60 30 — — — — — — — — — — 40 — B 10 20 10 — 100— — — — — — 22 — — — C 10 20 10 — — — 100 — — — — 22 — — — D 10 20 10 —100 — — — — — — 20 — — 2 E 10 20 10 10 100 — — — — — — 18 — — — F — 1010 10 100 — — — — 10 — 18 — — — G 10 20 10 — 100 — — — — — — 15 — — — H10 20 10 — 100 — — — — — — 15 — — 2 I 10 20 10 — — 100 — — — — — 18 — —— J 10 20 15 — — — — 100 — — — 20 — — — K 10 40 10 — — — — 100 — 10 — — 5 — — L 10 40 10 — — — — 100 — — 10 — 10 — — M 10 4 15 — — — 100 — — —— 15 — — — N 10 20 10 — — — 100 — — — — 22 — — — O 10 40 8 — — — — — 100— 10 —  5 — — P 10 40 10 — — — — — 100 — 10 — — — — Q 10 40 10 — — — — —100 — 10 —  5 — — R 10 40 10 — — — — 100 — — — — 10 — — S 10 40 20 — — —— 100 — — — — — — — T 10 40 20 — — — — 100 — — — — — — — U 10 20 — — —100 — — — — — 18 10 — — V 10 40 20 — — — — 100 — — — 20 — — — W  0 20 10— 100 — — — — — 10 18 — — — X — 67.5 35 13.75 — — — — — 17.5 — — — — —

EXAMPLE 7 Use of Zinc-Chelated 8-Hydroxyquinoline Solutions to TreatLesions in Humans

A solution corresponding to Sample B from Table 1 in Example 6 wasapplied topically to treat cancerous lesions, precancerous lesions, andwarts on human patients. Treatment included one to three single topicalapplications of the ointment, and was successful in every case. Apreferred treatment modality includes scraping the surface of epitheliallesions prior to application of the ointment, and this treatment isespecially beneficial in the treatment of hard warts. The lesion isinspected about two days after treatment. Additional ointment is appliedabout one week after the first application if necrosis of the lesion isnot apparent. In approximately one to two weeks time, the necroticlesion tissue is peeled away using forceps. Additional ointment isapplied to the wound margins to assure elimination of cancer cells andother diseased cells. Table 2 below lists the types of lesions that weresuccessfully treated, as well as the number of lesions that weresuccessfully treated. The total elapsed time from the topicalapplication to sloughing of the diseased tissue averaged approximatelytwelve days in each case. The patients who underwent treatment forvenereal warts reported much less discomfort than normally exists forchemical removal of venereal warts.

TABLE 2 SUCCESSFUL USE OF CHELATED 8-HYDROXYQUINOLINE IN HUMANS Numberof Lesion Type Lesions Location Basal cell 2 Left side of nose 1 Rightear 7 Right forehead 2 Forehead 1 Left Cheek 3 Scalp 1 Neck 3 Face 2Hands 1 Head 2 Left mandibular area 1 Arm 2 Left Temple 4 (Unspecified)Solar Keratosis 2 Head 1 Right forearm Human papilloma virus 1 Leftchest 1 (Unspecified) Venereal warts 2 Penis 1 (Unspecified) Maleveruoca warts 1 (Unspecificed) Kaposi's sarcoma 1 Arm 1 Leg 1 Wrist 1Foot

EXAMPLE 8 Use of Zinc-Chelated 8-Hydroxyquinoline Solutions to TreatLesions in Animals

A solution corresponding to sample B from Table 1 in Example 6 wasapplied topically to treat cancerous lesions and precancerous lesions onanimals. Treatment was successful in every case, with the exception ofone cat that died of sarcoma before the treatment was concluded. Table 3below lists the types of lesions that were successfully treated and thetype of animal on which treatment was successful.

TABLE 3 SUCCESSFUL USE OF CHELATED 8-HYDROXYQUINOLINE IN ANIMALS LesionType Type of Animal Eye cancer 3 Cows Squamous cell carcinoma 1 Horse(Penis) Sarcoid 10 Horses; 1 Mule Sarcoma 1 Dog (leg) Malignant melanoma2 Horses Rectal adenoma 1 Dog Sebaceous gland adenoma 1 Dog Histiocytoma1 Dog Sebaceous adenoma 1 Dog (two applications) Cutaneous Lymphoma 1Cat Sarcoma 2 Cats (one died before end of treatment); 1 Dog; 1 Bear

EXAMPLE 9 Titration of Zinc-Chelated 8-Hydroxyquinoline Solution toReduce Escharotic Effects While Optimizing Therapeutic Effect

Three samples of ointment were prepared corresponding to Sample B fromTable 1, except the lecithin/palmitate was not added and the weightpercentage amount of zinc chloride comprised 20%, 40%, 55% and 75%,respectively, for the samples. The solutions were each applied to apopulation of twenty five athymic nude mice with explanted lung cancerin the manner of Example 1. The 55% and 75% samples producedsubstantially immediate chemical escharotic effects. The 40% sampleproduced heightened redness proximate the lesion site, and wastherapeutically effective. At a 40% concentration, the ointment has theeffect of selectively eating away the lesion before the underlyingtissue has opportunity to regenerate. The 20% sample was therapeuticallyeffective without burning, and the underlying tissue has a chance toregenerate without the formation of an eschar, i.e., the 20% solutiondoes not tend to leave a hole at the lesion site, rather, no eschar isproduced because the necrotic lesion tissue remains in place while theunderlying tissue at least partially regenerates.

It is preferred not to use the Pluronic gels at high concentrations ofZnCl₂ because it has been observed that the gels degrade over time dueto an altered solution pH.

EXAMPLE 10 Use of Zinc-Chelated 8-Hydroxyquinoline on Human Patients ina Clinical Evaluation

Patient 1: A man entered a clinic to request treatment of a large growthon his right wrist. The growth was approximately eleven centimeters indiameter and circumscribed more than fifty percent of the wrist. Thesize and location of the growth meant that surgery would leave the manwith impaired wrist movement. The growth was diagnosed as basal cellcarcinoma. The surface of the carcinoma was scraped with a scalpel. Asolution corresponding to Sample B of Table 1 was applied to thecarcinoma using a cotton swab. The patient was subsequently monitoredfor two months. The patient complained of a burning sensation at thetime of initial application. The growth appeared red and inflamed at itsmargins on the first day after treatment. One week later, the growthappeared black and necrotic. Two weeks after treatment, the growth wasblack and necrotic to an extent permitting the dead growth to be peeledaway using forceps. A second application of solution B was made to thewound margins to assure that no cells from the excised cancer lesionhas\d escaped. Pathologic examination of tissue from the wound marginconfirmed that basal cell carcinoma was absent from healthy tissues atthe margin of the wound. Six weeks after treatment, the wound siteappeared as healthy tissue with no sign of basal cell carcinoma orscarring. Hair follicles were intact at the site of the former wound andbasal cell carcinoma. The. man retained full function in the affectedwrist.

Patient 2: A woman entered a clinic with a basal cell carcinomaapproximately one inch below the corner of her left eye. The growth wasapproximately one-half inch in diameter. The surgical prognosis was thatremoval of the growth would leave the woman with partial paralysis ofthe face and eye muscles. The surface of the carcinoma was scraped witha scalpel. A solution corresponding to solution B from Table 1 wasapplied to the carcinoma using a cotton swab. The patient wassubsequently monitored for two months. The patient complained of aburning sensation at the time of initial application. The growthappeared red and inflamed at its margins on the first day aftertreatment. One week later, the growth appeared black and necrotic. Twoweeks after treatment, the growth was black and necrotic permitting thecarcinoma to be peeled away by forceps. Pathologic examination of tissuefrom the wound margin confirmed that basal cell carcinoma was absentfrom healthy tissues at the margin of the wound. Pathologic examinationof tissue from the wound margin on the patient's face wrist confirmedthe absence of basal cell carcinoma. Six weeks after treatment, thewound site appeared as healthy tissue with no sign of basal cellcarcinoma or scarring. Hair follicles were intact at the site of theformer wound and basal cell carcinoma. The woman retained full functionin muscles of the face and eye.

Comparative trial: A patient had nodular pigmented lesions on his facesince childhood. Three of the tumors ulcerated. Diagnosis was nevoidbasal cell syndrome including multiple basal cell epitheliomas. Three ofthe tumors ulcerated. These were scraped and solution B from Table 1 wasapplied to these tumors. Necrotic tissue from the lesions was peeledaway by forceps two weeks later. Ten other tumors were removed bycryosurgery. None of these tumors recurred.

A second ointment was prepared to include the ingredients of solution B,but the oxyquinoline was absent. A third ointment was prepared toinclude the ingredients of solution B, but zinc chloride was absent. Thesecond and third ointments were applied to basal cell epitheliomas onthe patient's face. The second and third ointments were not effective inremoving the epitheliomas.

EXAMPLE 11 Veterinary Use of Zinc-Chelated 8-Hydroxyquinoline inClinical Evaluations

Female Domestic Short Hair Cat With Allergic Dermatitis: A female cataged six years developed a lesion approximately four inches long and twoinches wide in the flank ventral region. Antibiotic and surgicaltreatment modalities were tried without success at three differentanimal clinics. The lesion recurred within a few weeks after it wassurgically removed. The lesion was biopsied at the time of surgery. Thehistopathology report noted severe chronic (fibrotic) inflammation inthe upper dermis with plentiful mature mast cells and a moderately heavyscattering of other mixed leukocytes. The epidermis had occasionalshallow ulcers. A fungal stain tested negative for ringworm and surfaceyeasts, and no parasites were present. The diagnosis was hypersensitiveor allergic dermatitis.

The zinc chelated oxyquinoline formula of Example 1 was applied to coatthe lesion twice with the second application following the firstapplication by two weeks. The lesion was scraped prior to eachapplication. Two months and four days after the first application, thelesion had shrunk to the size of a pin head. A second biopsy confirmedthe existence of allergic dermatitis in this very small region.

Female Boston Terrier With Squamous Papilloma and Sebaceous GlandHyperplasia: An obese thirteen year old Boston terrier was diagnosedwith numerous squamous papilloma tumors on the face and one tumor on theright foot. The dog's owner desired a nonsurgical treatment due to thedog's age and obesity problems. A biopsy of lip tissue showed that theepithelium was markedly hyperblastic. The underlying growth containedlobules of cystic sebaceous glands. Some melanocytes and melanincontaining macrophages were observed.

Five tumors were treated by direct application to the tumor of the zincchelated oxyquinoline formula from Example 1. Application occurred onceper week, and three of the tumors were resolved after threeapplications. The two largest tumors included one lesion on the foreheadand one on the right foot. The tumor on the right foot resolved afterthe fourth treatment and the largest tumor on the forehead wassignificantly diminished in size. Treatment was discontinued when thedog's owner temporarily moved.

Kodiak Bear With Soft Tissue Fibrosarcoma: An eight year old male Kodiakbear was diagnosed with a tumor in the dorsolateral region proximate theright carpus. The bear was anaesthetized and the tumor was biopsied.Clinical diagnosis was that of myxoid fibrosarcoma with chondroiddifferentiation Grade 2. There was a low to medium chance of metastasis.The tumor measured 54.5 cm at the circumference, 17 cm proximal todistal, and 21 cm lateral to base of the tumor. A diagnostic cliniciannoted some bone invasion and recommended amputation of the afflictedlimb.

Study of the of the biopsy samples showed a moderately differentiatedtumor with different lines of differentiation varying from fibroustissue to early cartilage type tissue. Cells included spindle cellsforming bundles that intersected at various angles. Myxomatous andfibrous stroma were present. Nuclei were mildly pleomorphic, oval, andhad prominent nucleoli. Mitoses were occasionally present, and therewere occasional giant nuclear forms. In some areas, the cells werefibrous. In other areas, the cells were plump with myxoid material orearly embryonic cartilagenous type material. One area showed ananaplasia where the cells also had some giant nuclear forms andincreased mitotic rates. There were no normal tissue margins.

An injectable solution was prepared by mixing ten grams of the solutionQ from Table 1 with 100 grams of isopropyl alcohol. A 5 cc quantity often percent zinc 8-hydroxyquinoline in isopropyl alcohol was injectedinto the center of the tumor mass, and 3 cc were injected into each offour fingers of the tumor. The tumor was surgically removed two weekslater. Biopsy confirmed that the tumor was encapsulated at the time itwas removed. No bone invasion was seen at the time that the tumor wassurgically removed.

Mule With Sarcoid: An eight year old mule was diagnosed with a twelvemillimeter diameter sarcoid located between the ear and the left eye.Visual diagnosis was confirmed by biopsy. The lesion was scraped and thezinc chelated oxyquinoline formula of Example 1 was applied to cover thescraped lesion. The lesion disappeared within one month. Total remissionwas pronounced at a followup examination two months after treatment.

Gelding With Squamous Cell Carcinoma: A twenty-one year old gelding wasdiagnosed with squamous cell carcinoma located on the unpigmentedportions of the penis. Six lesions were present varying in diameter fromfive millimeters to twelve millimeters. The lesions were scraped andcovered with the zinc chelated oxyquinoline formula of Example 1.Application was repeated one week later with mild swelling observed onthe affected area. All lesions had completely healed at a followupexamination thirty days after the second treatment.

Feline Cutaneous Lymophoma: A cat had lesions on the pad areas of allfour feet. The lesions ranged in size from one half to two centimeters.Surgery to remove the lesions was impossible due to their locations andthe impairment of mobility due to surgical scarring. The lesions werescraped with a scalpel and solution B from Table 1 was applied to thelesions. The lesions sloughed off and the pad areas were completelyhealed within about four to six weeks.

EXAMPLE 12 Preparation of Orally Administrable Compositions

A solution was mixed to include equal 33 gram weights of8-hydroxyquinoline, sodium ascorbate, and zinc chloride in 90% ethanolwith sufficient water added to just dissolve the ingredients at 110° F.The mixture stood to ambient room temperature under a ventilation hoodfor twenty four hours. A vacuum aspirator was used to remove remainingliquid while coprecipitating the remaining moieties from solution. Theprecipitate was scraped from the beaker, supplemented with 5 grams ofascorbic acid, and ground to homogeneity with mortar and pestle to yieldan orally administrable composition.

An oral base may be substituted for any of the Pluronic, Aquabase, orPlastibase substances in Table 1 to provide orally administrablesolutions. The oral base solutions may be applied directly to lesions ofthe mouth, or they may be diluted for ingestion. The oral base is madeby mixing one gram of gelatin, two grams of sodiumcarboxymethylcellulose, one ml of isopropyl alcohol, fifty grams ofPlastibase, and 500 mg of xanthan gum. Forty seven grams of distilledwater are brought to a boil and added to the mixture, which stands toroom temperature. Plastibase is added to the cooled mixture until adesired gel consistency is achieved.

Those skilled in the art will understand that the preferred embodimentsdescribed above may be subjected to apparent modifications withoutdeparting from the true scope and spirit of the invention. Theinventors, accordingly, hereby state their intention to rely upon theDoctrine of Equivalents, in order to protect their full rights in theinvention.

1. A composition for use in treating epithelial lesions comprising:8-hydroxyquinoline; a metal chelating agent including a zinc salt; and acarrier, the zinc chloride and 8-hydroxyquinioline each being present inan amount ranging from five percent to forty percent of said compositionby weight, the composition being provided in a dosage formulation thatis suitable for administration against an epithelial lesion.
 2. Thecomposition as set forth in claim 1 wherein said therapeuticallyeffective amounts include a molar ratio between said 8-hydroxyquinolineand said metal chelating agent ranging from 1:1 to 1:N+1 and N is anoxidation state of metal in said metal chelating agent.
 3. Thecomposition as set forth in claim 2 wherein said ratio imparts a neutralcharge as 1:N.
 4. The composition as set forth in claim 2 wherein saidtherapeutically effective amounts include 8-hydroxyquinoline in anamount ranging from five percent to twenty percent by weight and N=2. 5.The composition as set forth in claim 1 wherein said carrier is a gelbase.
 6. The composition as set forth in claim 1 wherein said gel baseis a polyoxyalkylene ether derivative of propylene glycol.
 7. Thecomposition as set forth in claim 1 wherein said carrier contains apenetrant.
 8. The composition as set forth in claim 1 wherein saidpenetrant is selected from the group consisting of at least dimethylsulfoxide and lecithin.
 9. The composition as set forth in claim 1wherein said penetrant is dimethyl sulfoxide.
 10. The composition as setforth in claim 1 wherein said carrier contains an antioxidant.
 11. Thecomposition as set forth in claim 10 wherein said antioxidant isselected from the group consisting of at least nordihydroguiaretic acid,nordihydroguiaretic acid derivatives, and functional homologues ofnordihydroguiaretic acid.
 12. The composition as set forth in claim 10wherein said antioxidant is selected from a group consisting of ascorbicacid, and functional homologues of ascorbic acid.
 13. The composition asset forth in claim 1 wherein the therapeutically effective amounts areeffective against the lesions which are mammalian lesions furtherselected from the group consisting of venereal warts, male veruocawarts, lesions produced by human papilloma virus, basal cell carcinoma,solar keratosis, Kaposi's sarcoma, eye cancer, sarcoids, sarcoma,malignant melanoma, rectal adenoma, histiocytoma, and sebaceous adenoma.14. The composition of claim 1, wherein the dosage formulation is atopical formulation.
 15. The composition of claim 1, wherein the dosageformulation is an oral formulation.
 16. The composition of claim 1,wherein the dosage formulation is an injectable formulation.